Purpose: Nitric oxide-donating acetylsalicylic acid (NO-ASA) has been shown to possess an antineoplastic effect in Wnt-/b-catenin-active cancers. As chronic lymphocytic leukemia (CLL) cells exhibit aberrantly active Wnt signaling, we investigated the effect of the para-isomer of NO-ASA on CLL cell survival in vitro and in a CLL-like xenograft mouse model.Experimental Design: Apoptosis in primary CLL cells was determined by flow cytometric annexin V-FITC (fluorescein isothiocyanate)/PI (propidium iodide) staining and immunoblotting of caspases, poly(ADP-ribose) polymerase (PARP), and antiapoptotic proteins. Interference of NO-ASA with Wnt/ b-catenin signaling was analyzed through immunoblots of different pathway members. Influence of caspase activation was investigated by pretreatment with a pan-caspase inhibitor. CLL-like JVM3 cells were subcutaneously inoculated into irradiated nude mice that were treated with 100 mg of para-NO-ASA/kg of body weight p.o. (by mouth) for 21 days.Results: para-NO-ASA induced apoptosis in CLL cells with an LC 50 (lethal concentration) of 8.72 þ 0.04 mmol/L, whereas healthy blood cells were not affected. Furthermore, the compound induced caspase 9, caspase 3, and PARP cleavage. In addition, cleavage of b-catenin and downregulation of b-catenin/ lymphoid enhancer factor (Lef)-1 targets was observed. para-NO-ASA demonstrated strong antitumor efficacy in the xenograft mouse model with a tumor inhibtion rate of 83.4%. During therapy, no gross toxicity could be observed.Conclusions: para-NO-ASA selectively induces apoptosis in primary CLL cells and efficiently reduces tumor growthin a CLL-likexenograftmodel. As NO-ASA is orally available and is generally welltolerated, para-NO-ASA might be a promising new compound for CLL therapy. Clin Cancer Res; 17(2); 286-93. Ó2010 AACR.
No abstract
3451 Poster Board III-339 We and others have shown that vascular endothelial growth factor (VEGF) plays a pivotal role in growth, survival and migration of chronic lymphocytic leukemia (CLL). VEGF dependent survival benefits in CLL are thereby mediated via autocrine and paracrine loops. However, so far no approach was studied focusing on the selective inhibition of VEGF in CLL. Vatalanib (PTK787/ZK 222584) and pazopanib (GW786034) are potent orally available, selective VEGF tyrosine kinase inhibitors. The aim of the present investigation was i) to study the efficacy and selectivity of both inhibitors in CLL cells, ii) to simulate potential combination with conventional cytostatics in vitro and iii) to test the effect on CLL like tumor xenografts in a mouse model. Primary CLL as well as healthy B cells were incubated with varying concentrations of both inhibitors for different time periods. Cells were then treated with combinations of each inhibitor and fludarabine, vincristin and doxorubicin, respectively. Apoptosis induction was analysed by flow cytometry (annexin V-FITC/PI staining) and cell survival was additionally investigated by an ATP dependent fluorescence assay. Percentage of surviving cells was defined as double annexin/PI negative cells in flow cytometry analysis. For in vivo experiments, four-week old BALB/c nu/nu mice were grafted with cells of a human chronic B cell leukemia cell line (JVM3). After tumors reached a mean volume of 100 mm3 per group (10 mice), drugs were administered once daily by oral gavage at 100 mg/kg bodyweight. Tumor volume was measured every second day by calliper. Vatalanib and pazopanib effectively induced apoptosis in CLL cells in vitro in a dose and time dependent fashion. During 24h incubation of CLL cells vatalanib showed a 50% lethal concentration (LC50) of 46.7 μM (n=26) and pazopanib of 32.7μM (n=19). In contrast, survival of B cells derived from healthy donors was only slightly affected at high concentrations of both drugs, thereby suggesting a large therapeutic range. Healthy B cells survived 80.5 ± 4.5% after 24h incubation with 100 μM vatalanib. In contrast, CLL cells showed a survival of 27.8 ± 6.0% (n=5; p=0.0001). Survival of healthy B cells treated with 100 μM of pazopanib was 89.1 ± 2.2% whereas survival of CLL cells was only 40.8 ± 2.3% (n=5; p<0.0001). Combination of the low dosed drugs with conventional cytostatics like fludarabin, vincristin and doxorubicin showed synergistic effects and significantly increased apoptosis rates in vitro. Single drug treatment showed survival rates of 80.1 ± 5.7% for 10 μM vatalanib, 86.2 ± 10.1 for 10 μM fludarabin, 69.8 ± 6.1 % for 0.10 μM vincristin and 51.6 ± 5.2% for 10 μM of doxorubicin. The combination of 10 μM vatalanib with 10 μM fludarabin showed survival rates of 59.4 ± 11.6% (p=0.0167), with 0.1 μM vincristin 32.5 ± 7.2% (p=0.0095) and with doxorubicin 26.0 ± 11.8% (p=0.0381). Survival rates for single treatment with 10 μM pazopanib were 84.3 ± 4.3%. The combination of 10 μM pazopanib with 10 μM fludarabin showed survival rates of 55.0 ± 6.1% (p=0.0384), with 0.1 μM vincristin 42.8 ± 5.3 (p=0.0473) and with doxorubicin 41.5 ± 11.8% (n.s.). After three weeks of treatment of the xenograft mice with each inhibitor the mean tumor volume was 645.0 ± 241.2 mm3 with pazopanib (p=0.002), 671.8 ± 198.0 mm3 with vatalanib (p=0.002) and 2,458.3 ± 742.39 mm3 in the vehicle treated group. This translated into a mean tumor inhibition rate of 77.3% for pazopanib and 71.3% for vatalanib after 21 days of treatment. Summing up, specific inhibition of VEGF by vatalanib or pazopanib might be a promising new therapeutic approach in CLL. Both compounds are orally available and showed acceptable in vivo toxicities in clinical trials, promising good compliance for the treatment. Therefore, they are attractive candidates for further testing in CLL. Disclosures Hallek: BayerScheringAG: Honoraria, Research Funding.
<p>Supplementary Figures S1-S3; Supplementary Table S1.</p>
<div>Abstract<p><b>Purpose:</b> Nitric oxide–donating acetylsalicylic acid (NO-ASA) has been shown to possess an antineoplastic effect in Wnt-/β-catenin–active cancers. As chronic lymphocytic leukemia (CLL) cells exhibit aberrantly active Wnt signaling, we investigated the effect of the <i>para</i>-isomer of NO-ASA on CLL cell survival <i>in vitro</i> and in a CLL-like xenograft mouse model.</p><p><b>Experimental Design:</b> Apoptosis in primary CLL cells was determined by flow cytometric annexin V–FITC (fluorescein isothiocyanate)/PI (propidium iodide) staining and immunoblotting of caspases, poly(ADP-ribose) polymerase (PARP), and antiapoptotic proteins. Interference of NO-ASA with Wnt/β-catenin signaling was analyzed through immunoblots of different pathway members. Influence of caspase activation was investigated by pretreatment with a pan-caspase inhibitor. CLL-like JVM3 cells were subcutaneously inoculated into irradiated nude mice that were treated with 100 mg of <i>para</i>-NO-ASA/kg of body weight p.o. (by mouth) for 21 days.</p><p><b>Results:</b><i>para</i>-NO-ASA induced apoptosis in CLL cells with an LC<sub>50</sub> (lethal concentration) of 8.72 + 0.04 μmol/L, whereas healthy blood cells were not affected. Furthermore, the compound induced caspase 9, caspase 3, and PARP cleavage. In addition, cleavage of β-catenin and downregulation of β-catenin/lymphoid enhancer factor (Lef)–1 targets was observed. <i>para</i>-NO-ASA demonstrated strong antitumor efficacy in the xenograft mouse model with a tumor inhibtion rate of 83.4%. During therapy, no gross toxicity could be observed.</p><p><b>Conclusions:</b><i>para</i>-NO-ASA selectively induces apoptosis in primary CLL cells and efficiently reduces tumor growth in a CLL-like xenograft model. As NO-ASA is orally available and is generally well tolerated, <i>para</i>-NO-ASA might be a promising new compound for CLL therapy. <i>Clin Cancer Res; 17(2); 286–93. ©2010 AACR</i>.</p></div>
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.